The ability to differentiate between the various types of hepatic injury is of great significance in the treatment of both transplant patients and also individuals who suffer from other hepatic diseases which may affect the biliary system.
Glutathione S-transferases (GSTs) comprise a multigene family of proteins consisting mainly of alpha (.alpha.GST), mu (.mu.GST), pi (.pi.GST) and theta-class (.theta.GST) isoforms as defined by isoelectric point and are responsible for the detoxification of a range of xenobiotics, mainly via conjugation to glutathione (Beckett, G. J and Hayes, J. D., Advances in Clinical Chemistry (1993); 30, 281-380). Generally, the proteins are dimeric in nature consisting of two 25-27kDa subunits and may exist in homodimeric or heterodimeric forms. Pi Glutathione S-transferase (.pi.GST) is a homodimer, and is located in the cytoplasm of bile duct epithelial cells within the liver (Beckett G. J. and Hayes, J. D., (1993) supra). .alpha.GST is known to be present in hepatocytes within the liver and exists in both homodimeric and heterodimeric states (Campbell, J. A. H., et. al., Cancer (Philadelphia) (1991) 67, 1608-1613; Howie, A. F., et. al., Clin. Chem. Acta., (1988) 177, 65-76). This heterogenous GST distribution of .alpha. and .pi.GST suggests that the different isoenzymes have unique in vivo functions in different hepatic regions (Campbell, J. A. H., et. al., (1991) supra).
EP-A 0 640 145 discloses a method which assists in the early diagnosis of rejection in a liver transplant recipient and which comprises measuring an increase in plasma or serum .alpha.GST from the recipient in the absence of or preceding any change in plasma or serum transaminase. Thus, it has been conclusively demonstrated that measurement of the plasma .alpha.GST level facilitates monitoring of the post-transplant hepatic status by acting as an extremely sensitive, although not totally specific marker of graft rejection.
It is notable that .pi.GST has received no attention as a potential marker of graft rejection, a fact possibly due to the low levels of enzyme present in the biliary epithelial cells of the liver. There is some evidence, however, that .alpha. and .pi.GST are present in bile from both normal individuals and people suffering from specific cancers (e.g., cholangiocarcinoma) as measured by radio-immunoassay (Howie. A. F., et. al., Clin. Chem. Acta. (1989) 184, 269-278). Additionally, some authors have referenced the fact that measurement of serum and plasma .pi.GST levels may facilitate diagnosis of malignant tumours since .pi.GST appears to be specifically expressed in malignant tissue (Niitsu, Y., etal., Cancer (1989) 63, 317-323; Howie, A. F., et. al., Clin. Chem. (1990) 36(3), 453-456. and Hida, T., et al., Cancer (1994) 73(5), 1377-1382. None of the aforementioned authors allude to the fact that .pi.GST may have a role in the prediction of transplanted liver rejection or other liver/biliary disorders.
Since it is known that primary graft rejection generally occurs in the biliary tree within the liver (Ascher, N., (1993) In `Immunology of liver transplantation` Neuberger, J. and Adams, D. (eds)), it would appear that specific measurement of biliary or plasma .pi.GST levels may allow diagnosis of early rejection or facilitate discrimination between post-transplant hepatocellular or biliary damage. The importance of distinguishing between non-specific hepatic injury and graft rejection cannot be overstated since the treatment for each condition is entirely different. Furthermore, initiation of the incorrect treatment could be extremely deleterious to the health of an individual already severely ill. For example, if graft injury occurs due to viral infection (e.g., Hepatitis C re-infection or cytomegalovirus (CMV), it is necessary to carefully monitor the levels of anti-rejection immunosuppression treatment since excess immunosuppresive agents (e.g., cyclosporin A or FK506) would significantly impair the ability to fight viral infection. Conversely, failure to recognise genuine rejection from non-specific graft injury could lead to delay in augmentation of immunosuppressive therapy and ultimately lead to graft removal.
Accordingly, there is a need for methods of determining the hepatic status of an individual in various disease states or abnormal conditions of the liver.